CA1 pyramidal neurons of the hippocampus express various types of serotonin (5-HT) receptors, such as 5-HT(1A), 5-HT(4) and 5-HT(7) receptors, which couple to Galpha(i) or Galpha(s) proteins and operate on different intracellular signalling pathways. In the present paper we verify such differential serotonergic modulation for the hyperpolarization-activated current I(h). Activation of 5-HT(1A) receptors induced an augmentation of current-induced hyperpolarization responses, while the responses declined after 5-HT(4) receptors were activated. The resting potential of neurons hyperpolarized (-2.3 +/- 0.7 mV) after 5-HT(1A) receptor activation, activation of 5-HT(4) receptors depolarized neurons (+3.3 +/- 1.4 mV). Direct activation of adenylyl cyclase (AC) by forskolin also produced a depolarization. In voltage clamp, the Ih current was identified by its characteristic voltage- and time-dependency and by blockade with CsCl or ZD7288. Activation of 5-HT(1A) receptors reduced I(h) and shifted the activation curve to a more negative voltage by -5 mV at half-maximal activation. Activation of 5-HT(4) and 5-HT(7) receptors increased I(h) and shifted the activation curve to the right by +5 mV. Specific activation of 5-HT(4) receptors by BIMU8 increased membrane conductance and showed an increase in I(h) in a subset of cells, but did not induce a significant alteration in the activation curve. In order to verify spatial differences, we applied BIMU8 selectively to the soma and to the dendrites. Only somatic application induced receptor activation. These data are confirmed by immunofluorescence stainings with an antibody against the 5-HT(4) receptor, revealing receptor expression at the somata of the CA1 region. A similar expression pattern was found with a new antibody against 5-HT(7) receptors which reveals immunofluorescence staining on the cell bodies of pyramidal neurons.

Dinoflagellates are microbial eukaryotes that have exceptionally large nuclear genomes; however, their organelle genomes are small and fragmented and contain fewer genes than those of other eukaryotes. The genus Amoebophrya (Syndiniales) comprises endoparasites with high genetic diversity that can infect other dinoflagellates, such as those forming harmful algal blooms (e.g., Alexandrium). We sequenced the genome (~100 Mb) of Amoebophrya ceratii to investigate the early evolution of genomic characters in dinoflagellates. The A. ceratii genome encodes almost all essential biosynthetic pathways for self-sustaining cellular metabolism, suggesting a limited dependency on its host. Although dinoflagellates are thought to have descended from a photosynthetic ancestor, A. ceratii appears to have completely lost its plastid and nearly all genes of plastid origin. Functional mitochondria persist in all life stages of A. ceratii, but we found no evidence for the presence of a mitochondrial genome. Instead, all mitochondrial proteins appear to be lost or encoded in the A. ceratii nucleus.

Although the emergence of complex biomineralized forms has been investigated for over a century, still little is known on how single cells control morphology of skeletal structures, such as frustules, shells, spicules, or scales. We have run experiments on the shell formation in foraminifera, unicellular, mainly marine organisms that can build shells by successive additions of chambers. We used live imaging to discover that all stages of chamber/shell formation are controlled by dedicated actin-driven pseudopodial structures. Successive reorganization of an F-actin meshwork, associated with microtubular structures, is actively involved in formation of protective envelope, followed by dynamic scaffolding of chamber morphology. Then lamellar dynamic templates create a confined space and control mineralization separated from seawater. These observations exclude extracellular calcification assumed in selected foraminiferal clades, and instead suggest a semiintracellular biomineralization pattern known from other unicellular calcifying and silicifying organisms. These results give a challenging prospect to decipher the vital effect on geochemical proxies applied to paleoceanographic reconstructions. They have further implications for understanding multiscale complexity of biomineralization and show a prospect for material science applications.

We have reported recently that the mouse 5-hydroxytryptamine(4a) (5-HT 4(a) ) receptor undergoes dynamic palmitoylation (Ponimaskin, E. G., Schmidt, M. F., Heine, M., Bickmeyer, U., and Richter, D. W. (2001) Biochem. J. 353, 627-663). In the present study, conserved cysteine residues 328/329 in the carboxyl terminus of the 5-HT 4(a) receptor were identified as potential acylation sites. In contrast to other palmitoylated Gprotein-coupled receptors, the additional cysteine residue 386 positioned close to the COOH-terminal end of the receptor was also found to be palmitoylated. Using pulse and pulse-chase labeling techniques, we demonstrated that palmitoylation of individual cysteines is a reversible process and that agonist stimulation of the 5-HT 4(a) receptor independently increases the rate of palmitate turnover for both acylation sites. Analysis of acylation-deficient mutants revealed that non-palmitoylated 5-HT 4(a) receptors were indistinguishable from the wild type in their ability to interact with G s , to stimulate the adenylyl cyclase activity and to activate cyclic nucleotide-sensitive cation channels after agonist stimulation. The most distinctive finding of the present study was the ability of palmitoylation to modulate the agonist-independent constitutive 5-HT 4(a) receptor activity. We demonstrated that mutation of the proximal palmitoylation site (Cys 328 3 Ser/Cys 329 3 Ser) significantly increases the capacity of receptors to convert from the inactive (R) to the active (R*) form in the absence of agonist. In contrast, the rate of isomerization from R to R* for the Cys 386 3 Ser as well as for the triple, nonpalmitoylated mutant (Cys 328 3 Ser/Cys 329 3 Ser/Cys 386 3 Ser) was similar to that obtained for the wild type.Covalent binding of long chain saturated fatty acids occurs within a wide variety of cellular as well as viral polypeptides (1-3). Two of the most common modifications involve acylation with myristate (N-myristoylation) and palmitate (S-acylation). Myristic acid is usually attached co-translationally to the NH 2 -terminal glycine residue in an amide linkage by N-myristoyltransferase (4, 5). Palmitic acid is attached to cysteine residues via a labile thioester bond (6). In contrast to myristate, which generally remains attached to the polypeptides until protein degradation, palmitic acid is added post-translationally and turns over rapidly as a protein itself (7,8).Among the cellular palmitoylated proteins, polypeptides involved in signal transduction (e.g. receptors, G-protein ␣-subunits, and adenylyl cyclases) are prevalent. With the finding that palmitoylation states of several proteins may be dynamically regulated, it is now widely accepted that repeated cycles of palmitoylation and depalmitoylation could have important functional consequences for signaling (9 -11). In G-protein-coupled receptors (GPCRs), 1 the functions of palmitoylation cover the wide spectrum of their biological activities: from coupling to G-proteins and regulated endocytosis to receptor phosphorylation and d...

Thermal tolerance windows serve as a powerful tool for estimating the vulnerability of marine species and their life stages to increasing temperature means and extremes. However, it remains uncertain to which extent additional drivers, such as ocean acidification, modify organismal responses to temperature. This study investigated the effects of CO -driven ocean acidification on embryonic thermal sensitivity and performance in Atlantic cod, Gadus morhua, from the Kattegat. Fertilized eggs were exposed to factorial combinations of two PCO conditions (400 μatm vs. 1100 μatm) and five temperature treatments (0, 3, 6, 9 and 12 °C), which allow identifying both lower and upper thermal tolerance thresholds. We quantified hatching success, oxygen consumption (MO ) and mitochondrial functioning of embryos as well as larval morphometrics at hatch and the abundance of acid-base-relevant ionocytes on the yolk sac epithelium of newly hatched larvae. Hatching success was high under ambient spawning conditions (3-6 °C), but decreased towards both cold and warm temperature extremes. Elevated PCO caused a significant decrease in hatching success, particularly at cold (3 and 0 °C) and warm (12 °C) temperatures. Warming imposed limitations to MO and mitochondrial capacities. Elevated PCO stimulated MO at cold and intermediate temperatures, but exacerbated warming-induced constraints on MO , indicating a synergistic interaction with temperature. Mitochondrial functioning was not affected by PCO . Increased MO in response to elevated PCO was paralleled by reduced larval size at hatch. Finally, ionocyte abundance decreased with increasing temperature, but did not differ between PCO treatments. Our results demonstrate increased thermal sensitivity of cod embryos under future PCO conditions and suggest that acclimation to elevated PCO requires reallocation of limited resources at the expense of embryonic growth. We conclude that ocean acidification constrains the thermal performance window of embryos, which has important implication for the susceptibility of cod to projected climate change.